The goal of this project is to identify factors that regulate secretion of neuropeptides generally, with a specific focus on insulin-like growth factors. The motivation for this project is two-fold. First, insulin secretion, and its misregulation, plays a pivotal role in aging, diabetes, and obesity. Second, while a great deal has been learned about mechanisms regulating secretion of classical neurotransmitters, far less is known about those regulating secretion of neuropeptides and hormones. Classical neurotransmitters are packaged in synaptic vesicles (SVs), which are clustered at active zones. Neuropeptides are packaged into large dense core vesicles (DCVs), and are distributed throughout axons and dendrites. Secretion of SVs occurs at active zones, in a rapid, phasic manner in response to single action potentials. Secretion of DCVs occurs typically after trains of depolarization, fusion events occur far from active zones, and they occur relatively slowly following depolarization. Following exocytosis, the SV pool is rapidly reconstituted at nerve terminals by endocytic recycling of SV components, and refilling with neurotransmitters. By contrast, the releasable pool of DCVs must be reconstituted by anterograde transport of immature secretory granules from the soma. Relatively little is known about the biochemical basis for these differences. We propose to identify factors that are required for or that regulate DCV secretion, using C. elegans as a model system. In summary, changes in insulin secretion have profound effects on human health. These studies should provide new insights into the cellular mechanisms regulating secretion of insulin and other neuropeptides. First, we will screen genes that are known to be required for SV secretion to determine which are also required for DCV biogenesis, trafficking, or secretion. Second, using RNAi, we will screen a large set of genes (180) required for neuromuscular signaling for effects on DCV biogenesis, trafficking, or secretion. Third, we will characterize genes and physiological conditions that regulate insulin secretion, and determine the impact of these pathways on lifespan and metabolism. Finally, we will screen known and candidate targets of insulin signaling for defects in synaptic transmission. These genes should provide insights into the mechanisms by which insulin regulates synaptic transmission and behavior. In summary, changes in insulin secretion have profound effects on human health. These studies should provide new insights into the cellular mechanisms regulating secretion of insulin and other neuropeptides.